The long-term objective of this proposal is to develop a safe and efficacious AIDS vaccine. A number of candidate AIDS vaccines derived from the attenuated poxvirus Modified Vaccinia virus Ankara (MVA) have been (or currently are being) evaluated in clinical trials. However, a number of factors limit the immunogenicity and utility of these vaccine candidates. These include priming HIV-specific T cell responses that are of limited breadth, due to unfavorable antigenic competition with poxvirus proteins encoded by the vector, as well as the development of potent anti-vector immunity, following immunization with recombinant MVA vaccines, that progressively diminishes the effectiveness of homologous booster immunizations to boost HIV-specific T and B cell responses. As a result, and in light of the STEP trial results, there is an urgent need to discover new viral vector modifications and immunization strategies that maximize the magnitude, quality, and breadth of HIV-specific T cell responses that are elicited by vaccine vectors. In addition, it is quite likely that an AIDS vaccine will need to engender broadly neutralizing antibody responses, in addition to CTLs, to be maximally effective. As such, it is imperative to identify the best vectors and methodologies for presenting relevant envelope antigens (once developed) in order to elicit high-titer neutralizing antibody responses that are durable. Toward achieving these goals, we propose to evaluate a number of novel vector modifications that are hypothesized to enhance the cellular and humoral immunogenicity of MVA-based AIDS vaccines, and to mitigate the negative effects of pre-existing, or immunization-induced, vector-specific immunity. We will pursue the following specific aims: 1) We will test the hypothesis that the immunogenicity of recombinant MVA-based AIDS vaccines, particularly in hosts exhibiting vector-specific immunity, can be significantly enhanced through vector modifications that delete relevant determinants of MVA-specific humoral immunity and that attenuate the ability of complement to neutralize virion infectivity or to facilitate the clearance of MVA-transduced cells in vivo;2) We will test the hypothesis that the breadth and magnitude of HIV-specific T cell and antibody responses that are elicited by recombinant MVA-based AIDS vaccines can be significantly enhanced through vector modifications that specifically target antigenic stimulation of TLR-5-mediated innate immune signaling pathways. PUBLIC HEALTH RELEVANCE: The world desperately needs an AIDS vaccine. We propose to develop novel Modified Vaccinia Ankara (MVA)-based AIDS vaccines that are more immunogenic than vectors derived from the parental strain of MVA. Rationally improving MVA vectors to elicit higher levels of more highly diverse HIV-specific T cell and antibody responses and to mitigate their induction of, and sensitivity to, vector-specific neutralizing antibodies, should result in new candidate AIDS vaccines that exhibit greater efficacy than current alternatives.